This study examined the antigenic and genetic diversity of serogroup B Neisseria meningitidis (MenB) recovered from invasive meningococcal disease (IMD) cases in Ontario, Canada for the 10 year period of 2001 to 2010 during which no MenB outbreaks had occurred. MenB was found to be responsible for 39% of all IMD cases, with the remaining cases mainly caused by serogroup Y (28%), serogroup C (24%), and serogroup W135 (8%). One hundred and ninety-three individual MenB case isolates were collected and characterized. Of the 88 sequence types (STs) identified, 75 were grouped into 14 known clonal complexes (CCs) while 13 STs were not assigned to any known CC. Fifty-seven different PorA genotypes and 88 STs defined the diversity of invasive MenB in Ontario, which supported the endemic nature of MenB disease in Ontario. Despite the presence of the hyper-virulent ST-41/44 and ST-32 CCs, no single ST was predominant and responsible for a large number of IMD cases. Although the Québec outbreak clone of ST-269 was also found in Ontario, the 20 case isolates were genetically diverse, grouped into 7 STs and without a predominant PorA genotype. eBURST analysis identified a new CC responsible for 14.5% of the MenB case isolates. The 6 most common PorA VR2 genotypes (VR2-9, 4, 14, 16, 13-1, and 16-3) were found in 67% of invasive MenB isolates.